EP3059299B1 - Zellkultivierungsvorrichtung - Google Patents

Zellkultivierungsvorrichtung Download PDF

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Publication number
EP3059299B1
EP3059299B1 EP13895619.8A EP13895619A EP3059299B1 EP 3059299 B1 EP3059299 B1 EP 3059299B1 EP 13895619 A EP13895619 A EP 13895619A EP 3059299 B1 EP3059299 B1 EP 3059299B1
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EP
European Patent Office
Prior art keywords
vessel
liquid
culture
valve
culture medium
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EP13895619.8A
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English (en)
French (fr)
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EP3059299A1 (de
EP3059299A4 (de
Inventor
Masaharu Kiyama
Guangbin ZHOU
Takayuki Nozaki
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Hitachi Ltd
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Hitachi Ltd
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Publication of EP3059299A4 publication Critical patent/EP3059299A4/de
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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M41/00—Means for regulation, monitoring, measurement or control, e.g. flow regulation
    • C12M41/30—Means for regulation, monitoring, measurement or control, e.g. flow regulation of concentration
    • C12M41/34—Means for regulation, monitoring, measurement or control, e.g. flow regulation of concentration of gas
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M29/00—Means for introduction, extraction or recirculation of materials, e.g. pumps
    • C12M29/26—Conditioning fluids entering or exiting the reaction vessel

Definitions

  • the present invention relates to an automatic cell culture system that cultures cells and a liquid sending device.
  • Fig. 4 is a diagram of a ventilation method for the culture vessel that conducts two-layer culture in the configurations in Figs. 2 and 3 .
  • a mixed gas cylinder 17 containing CO 2 and O 2 is connected to a sparger bottle 19.
  • the CO 2 gas sent from the cylinder 17 is passed into pure water 18 in the sparger bottle 19, and the gas is humidified at the optimum humidity, and ready for use.
  • a liquid culture medium that has not reached the vessel remains in the liquid sending tube through which a liquid culture medium has been sent.
  • the gaseous phase then moves to send the remaining liquid culture medium, and the liquid culture medium reaches the trap bottle 24.
  • the liquid having passed through the inside of the liquid sending tube and reached the trap bottle 24 is condensed at the vessel side opening 26, and then dropped onto the inner bottom part of the bottle.
  • the gaseous phase passes through the atmosphere side opening 27 communicating inside the bottle, reaches the air communication filter 29, and then is released to the atmosphere.
  • the pumps mean roller pumps.
  • the filters described above are a filter that takes in a gas externally from the flow channel or discharges a gas for adjusting the atmospheric pressure in the inside of the flow channel.
  • filters having quality of blocking particles in particle size of 0.22 ⁇ m or greater, bacterial cells, and viruses are used.
  • the filters mean similar ones.
  • the first culture vessel 55 is a hermetical vessel including a main body 59 and a lid 60.
  • the first culture vessel 55 can hold a second vessel 61 and a first vessel 62 on the inner bottom part of the main body 59.
  • the second vessel 61 can hold and culture cells.
  • the first vessel 62 can hold and culture cells.
  • the culture surface of the first vessel 62 is formed of a substance permeable membrane.
  • the first culture vessel 55 is a culture vessel that conducts a so-called co-culture, which is a culture method in which the membrane transmits only growth factors produced from vegetative cells cultured in the second vessel 61 for promoting the growth of cells cultured in the first vessel.
  • the main body 59, the lid 60, and the first port 63 to the fourth port 66 were prepared by injection molding with polycarbonate for their materials.
  • a cell culture insert (six wells), model No. 353090, made by Becton, Dickinson and Company, was used.
  • the substance permeable membrane 9 was polymerized with N-isopropylacrylamide, which is a temperature responsive polymer monomer, by electron beams. Thus, a temperature responsive culture surface was prepared.
  • a 35-mm cell culture surface processed dish, model No. 430165, made by Corning Incorporated was used.
  • an incubator For the thermostat, an incubator (Toyo Engineering Works, Ltd., model No. TVHA60WA12A) was used. The chamber temperature of the thermostat was operated at a temperature of 37°C.
  • an electronic low temperature chamber For the refrigerator unit, an electronic low temperature chamber (Toyo Engineering Works, Ltd., model No. THS030PA) was used. The chamber temperature of the refrigerator unit was operated at a temperature of 4°C.
  • Holding the trap bottle at a temperature lower than a room temperature improves the efficiency of condensing a liquid from the gaseous phase containing the liquid.
  • the liquid phase is prevented from reaching the air communication filter, and the filter is prevented from being clogged. Consequently, the internal pressure of the culture vessel can be maintained at a normal pressure.

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  • Chemical & Material Sciences (AREA)
  • Wood Science & Technology (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Health & Medical Sciences (AREA)
  • Organic Chemistry (AREA)
  • Zoology (AREA)
  • Biomedical Technology (AREA)
  • Sustainable Development (AREA)
  • Microbiology (AREA)
  • Biochemistry (AREA)
  • General Engineering & Computer Science (AREA)
  • General Health & Medical Sciences (AREA)
  • Genetics & Genomics (AREA)
  • Biotechnology (AREA)
  • Analytical Chemistry (AREA)
  • Apparatus Associated With Microorganisms And Enzymes (AREA)

Claims (1)

  1. Automatisches Zellkultursystem (31),
    das einen Kulturbehälter (1, 55, 57) zum Durchführen einer Zweischichtkultur, der einen ersten Behälter (2, 62) und einen zweiten Behälter (3, 61) umfasst, und eine Flüssigkeitsleitungs- und -auslasseinheit im Kulturbehälter umfasst;
    wobei der Kulturbehälter (1, 55, 57) in einem Thermostaten (32) angeordnet ist und
    wobei im automatischen Zellkultursystem (31):
    der erste Behälter (2) mit einem Flüssigkeitsleitungsrohr (4), das konfiguriert ist, dem ersten Behälter (2) ein flüssiges Kulturmedium hinzuzufügen, und mit einem Auslassrohr (5), das konfiguriert ist, das flüssige Kulturmedium aus dem ersten Behälter (2) auszulassen, versehen ist;
    der zweite Behälter (3) mit einem Flüssigkeitsleitungsrohr (6), das konfiguriert ist, dem zweiten Behälter (3) ein flüssiges Kulturmedium hinzuzufügen, und mit einem Auslassrohr (7), das konfiguriert ist, das flüssige Kulturmedium aus dem zweiten Behälter (3) auszulassen, versehen ist, und
    das Flüssigkeitsleitungsrohr (4), das konfiguriert ist, dem ersten Behälter (2) das flüssige Kulturmedium hinzuzufügen, durch eine erste Pumpe (8) und eine Flüssigkeits-Ein/Aus-Ventil (16) mit einer Flüssigkeitsflasche (9) verbunden ist, die konfiguriert ist, ein flüssiges Kulturmedium zu halten, das zum ersten Behälter (2) geleitet werden soll; wobei das Flüssigkeitsleitungsrohr (6), das konfiguriert ist, dem zweiten Behälter (3) ein flüssiges Kulturmedium hinzuzufügen, durch eine zweite Pumpe (10) mit der Flüssigkeitsflasche (9) verbunden ist, die konfiguriert ist, ein flüssiges Kulturmedium zu halten, das zum zweiten Behälter (3) geleitet werden soll;
    das Flüssigkeitsleitungsrohr (6) zwischen der zweiten Pumpe (10) und einem Punkt in der Mitte des Weges zum Kulturbehälter (1) in zwei Kanäle verzweigt ist und konfiguriert ist, durch ein Auslass-Ein/Aus-Ventil (14) und einen Luftverbindungsfilter (15) als ein Auslassrohr (13) zu einer Atmosphäre zu öffnen;
    eine dritte Pumpe (11) konfiguriert ist, das flüssige Kulturmedium aus dem Auslassrohr (5) zu einer Rückgewinnungsflasche auszulassen; und
    eine vierte Pumpe (12) konfiguriert ist, das flüssige Kulturmedium aus dem Auslassrohr (5) zu einer weiteren Rückgewinnungsflasche auszulassen;
    und wobei:
    der erste Behälter (2, 62) und der zweite Behälter (3, 61) jeweils Folgendes umfassen:
    - einen ersten Anschluss (63, 65), der konfiguriert ist, eine Flüssigkeit oder ein Gas dem jeweiligen ersten (2, 62) oder zweiten Behälter (3, 61) zuzuführen und ein Gas aus dem Kulturbehälter (1, 55, 57) auszulassen;
    - einen zweiten Anschluss (64, 66), der konfiguriert ist, eine Flüssigkeit auszulassen, die sich im Inneren des jeweiligen ersten (2, 62) oder zweiten Behälters (3, 61) befindet;
    wobei der Kulturbehälter (1, 55, 57) ferner Folgendes umfasst:
    - einen Strömungskanal, der konfiguriert ist, das Gas, das aus dem ersten Anschluss (63, 65) des jeweiligen ersten (2, 62) oder zweiten Behälters (3, 61) ausgelassen wird, mit einem Atmosphärendruck zur Außenluft auszulassen;
    - den Luftverbindungsfilter (15, 29, 52), der auf dem Strömungskanal vorgesehen ist; und
    - eine Abscheideeinheit, die zwischen dem ersten Anschluss (63) und dem Luftverbindungsfilter (15, 29, 52) auf dem Strömungskanal vorgesehen ist, wobei die Abscheideeinheit konfiguriert ist, Feuchtigkeit in dem ausgelassenen Gas abzuscheiden, und
    wobei:
    das Flüssigkeitsleitungsrohr (4) ferner durch die erste Pumpe (8) und das Flüssigkeits-Ein/Aus-Ventil (16) mit der Flüssigkeitsflasche (9) verbunden ist, die konfiguriert ist, ein flüssiges Kulturmedium zu halten, das zum ersten und zweiten Behälter (2) geleitet werden soll; und
    das Flüssigkeitsleitungsrohr (6) durch die zweite Pumpe (10) mit der Flüssigkeitsflasche (9) verbunden ist, die konfiguriert ist, das flüssige Kulturmedium zu halten, das zum zweiten Behälter (3) geleitet werden soll, derart, dass dann, wenn das Flüssigkeits-Ein/Aus-Ventil (16) und das Auslass-Ein/Aus-Ventil (14) und die erste Pumpe (8) betrieben werden, eine Flüssigkeit sich aus der Flüssigkeitsflasche (9) bewegt, durch das Flüssigkeitsleitungsrohr (4) hindurchtritt und das Innere des ersten Behälters (2) des Kulturbehälters (1) erreicht, und
    dann, wenn eine dritte Pumpe (11) und eine vierte Pumpe (12) zum Auslassen nicht betrieben werden, das Flüssigkeitsleitungsrohr (6) des zweiten Behälters (3) als ein Belüftungsrohr verwendet werden soll, indem der Kulturbehälter (1) verschlossen wird und dieser luftdicht verschlossen gehalten wird, derart, dass eine gasförmige Phase des Kulturbehälters (1) entlang des Strömungskanals durch das Flüssigkeitsleitungsrohr (6) durch das geöffnete Auslass-Ein/Aus-Ventil (14) hindurchströmt und mit der Atmosphäre in Verbindung steht.
EP13895619.8A 2013-10-15 2013-10-15 Zellkultivierungsvorrichtung Active EP3059299B1 (de)

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
PCT/JP2013/077987 WO2015056302A1 (ja) 2013-10-15 2013-10-15 細胞培養装置

Publications (3)

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EP3059299A1 EP3059299A1 (de) 2016-08-24
EP3059299A4 EP3059299A4 (de) 2017-06-07
EP3059299B1 true EP3059299B1 (de) 2020-05-13

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US (1) US10428300B2 (de)
EP (1) EP3059299B1 (de)
JP (1) JP6279597B2 (de)
WO (1) WO2015056302A1 (de)

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CN108165603A (zh) 2013-01-25 2018-06-15 艾克斯赛尔生物科学公司 用于选择性富集靶细胞的方法、组合物、试剂盒及系统
WO2016013070A1 (ja) 2014-07-23 2016-01-28 株式会社日立製作所 送液装置、及び細胞培養装置
WO2017163326A1 (ja) * 2016-03-23 2017-09-28 株式会社日立製作所 培養容器及びそれを用いた自動培養装置
WO2019155033A1 (en) * 2018-02-09 2019-08-15 General Electric Company Bioprocessing vessel
US11920119B2 (en) 2018-02-09 2024-03-05 Global Life Sciences Solutions Usa Llc Systems and methods for bioprocessing
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CN108441424B (zh) * 2018-04-23 2023-08-22 福州大北农生物技术有限公司 一种微载体生物反应罐及其培养猪圆环病毒2型方法
CN108315256B (zh) * 2018-04-26 2024-04-09 上海久博生物工程有限公司 活性通气组件及其活性通气式生物反应器和细胞培养器
EP3819370A4 (de) * 2018-07-05 2021-08-18 FUJIFILM Corporation Zellzüchtungsvorrichtung und rührverfahren
CN113015429A (zh) * 2018-10-02 2021-06-22 学校法人同志社 用于保存角膜内皮细胞的方法和容器
CN110938544A (zh) * 2019-12-02 2020-03-31 宇环智能科技(徐州)有限公司 一种生物技术用细胞培养装置
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Also Published As

Publication number Publication date
WO2015056302A1 (ja) 2015-04-23
EP3059299A1 (de) 2016-08-24
EP3059299A4 (de) 2017-06-07
US10428300B2 (en) 2019-10-01
JPWO2015056302A1 (ja) 2017-03-09
JP6279597B2 (ja) 2018-02-14
US20160244711A1 (en) 2016-08-25

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